| Description | E1373491Component50 T100 T200 TStorage conditionsQuantity Per TestE1373491AEdU(10 mM)100 µL200 µL400 µL-20℃.Store in the dark.2 µL per 1.0-2.0 × 10⁶ cellsE1373491BAF488 azide100 µL200 µL400 µL-20℃.Store in the dark.2 µL&E1373491Component50 T100 T200 TStorage conditionsQuantity Per TestE1373491AEdU(10 mM)100 µL200 µL400 µL-20℃.Store in the dark.2 µL per 1.0-2.0 × 10⁶ cellsE1373491BAF488 azide100 µL200 µL400 µL-20℃.Store in the dark.2 µL per 1.0-2.0 × 10⁶ cellsE1373491CClick Reaction Buffer13 mL26 mL52 mL-20℃.Store in the dark.243 µL per 1.0-2.0 × 10⁶ cellsE1373491DCuSO4250 µL500 µL1000 µL-20℃.5 µL per 1.0-2.0 × 10⁶ cellsE1373491EClick Additive248 mg496 mg992 mg-20℃.Store in the dark.250 µL per 1.0-2.0 × 10⁶ cellsE1373491FDAPI Staining Solution（1000×）25 µL50 µL100 µL-20℃.Store in the dark.0.5 µL per 1.0-2.0 × 10⁶ cellsProduct IntroductionCell proliferation assays are widely used in the evaluation of cell viability, genotoxicity, and the efficacy of antitumor drugs. Direct detection of DNA synthesis in cells is considered the most accurate method for assessing cell proliferation. EdU (5-ethynyl-2′-deoxyuridine) is a novel thymidine (thymine deoxyribonucleoside) analogue. During DNA synthesis, EdU can be incorporated into newly synthesized DNA in place of thymidine. The ethynyl group on EdU can undergo a covalent reaction with fluorescently labeled small-molecule azide probes (such as Azide Alexa Fluor 488, Azide Alexa Fluor 555, Azide Alexa Fluor 594, Azide Alexa Fluor 647, etc.) via Cu(I)-catalyzed click chemistry, forming a stable triazole ring. This reaction is highly efficient and is referred to as the Click reaction. Through this process, newly synthesized DNA is labeled with the corresponding fluorescent probes, enabling the detection of proliferating cells using appropriate fluorescence detection equipment. Usage Protocol1. Preparation1) Preparation of Click Additive Solution: For a 50-test kit: Add 12.5 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. For a 100-test kit: Add 25 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. For a 200-test kit: Add 50 mL of pre-chilled deionized water to the tube. Mix thoroughly until completely dissolved to obtain the Click Additive Solution. After preparation, aliquot the solution as needed and store at -20°C. If a white precipitate forms after dissolution, invert the tube repeatedly until it is fully dissolved before use. If the solution turns brown, it indicates degradation of the active component; discard it.2) Upon initial dissolution of the Click Reaction Buffer, aliquot it according to the number of samples per experiment and store at -20°C.2. EdU Labeling of CellsIt is recommended to use a final EdU concentration of 10 µM (1×). A 1:500 dilution of EdU (10 mM) in cell culture medium yields a 2× EdU working solution (20 µM). Mix an equal volume of pre-warmed (37°C) 2× EdU working solution (20 µM) with the cell suspension to achieve a final 1× EdU concentration. Incubate in a 37°C, 5% CO₂ incubator. Factors such as cell culture medium, cell density, cell type, and other experimental conditions may affect labeling efficiency. Therefore, the optimal EdU concentration and labeling duration must be empirically determined based on the cell type under investigation.3. Fixation and Permeabilization1) Harvest cells and centrifuge at 300 ×g for 5 min. Wash cells twice with PBS containing 2% FBS.2) Fix cells with 4% paraformaldehyde solution. Mix thoroughly and incubate for 15 min at room temperature protected from light.3) Collect cells and centrifuge at 300 × g for 5 min. Wash cells twice.4) Resuspend cells in PBS containing 0.1% Triton X-100. Mix well and incubate for 15 min at room temperature.5) Centrifuge at 300 × g for 5 min and wash cells twice.4. Fluorescent Labeling1) This protocol is based on a 500 µL reaction system per 2 × 10⁶ cells. The volume of the Click reaction mixture can be adjusted according to the experimental sample size.2) Centrifuge the cells at 300 ×g for 5 minutes. Add 500 µL of Click reaction mixture per sample, mix gently, and incubate for 30 minutes at room temperature protected from light.3) After the reaction, wash the cells twice with PBS containing 2% FBS.4) Dilute the DAPI Staining Solution (1000×) to 1× using PBS containing 2% FBS. Add 250 µL of the diluted DAPI solution to each sample and incubate for 5 minutes at room temperature.5) Add an additional 250 µL of PBS containing 2% FBS, mix gently, and proceed to detection using an appropriate flow cytometry instrumentPrecautions1. Strictly adhere to the component order and volumes specified in the table above when preparing the Click reaction mixture, as deviations may affect subsequent experimental results.2. The Click reaction mixture must be used within 15 minutes of preparation.3. To avoid fluorescence quenching, perform detection as soon as possible after sample preparation... Read More | Description：Acetylcholinesterases (AChEs) are enzymes that hydrolyze the neurotransmitter acetylcholine (ACh) to acetate and choline. AChE is located at the synaptic cleft and functions to terminate synaptic transmission by catalyzing the breakdown of ACh allowing cholinergic neurons to Description：Acetylcholinesterases (AChEs) are enzymes that hydrolyze the neurotransmitter acetylcholine (ACh) to acetate and choline. AChE is located at the synaptic cleft and functions to terminate synaptic transmission by catalyzing the breakdown of ACh allowing cholinergic neurons to return to a resting state after activation. It is also found in membranes of red blood cells, motor and sensory fibers, muscles, nerves and central and peripheral tissues. Changes in AChE activity may result from exposure to certain insecticides, which act as cholinesterase inhibitors. Inhibitors of AChE are also used to treat certain conditions such as dementia.Acetylcholinesterase activity assay kit has been used to determine the activity of acetylcholinesterase in a rat organophosphate model and in brain tissue homogenates.Principle：Acetylcholinesterase can catalyze the hydrolysis of acetylcholine to choline, and the reaction of choline with disulfide p-nitrobenzoic acid to produce 5-merhydryl-nitrobenzoic acid (TNB). The product has a characteristic absorption peak at 412 nm, and the activity of acetylcholinesterase can be characterized by the change of light absorption valueThe Dilution Calculator EquationConcentration (start)xVolume (start)= Concentration (final)× Volume (final)This equation is commonly abbreviated as: C1V1 = C2V2... Read More | Product IntroductionAlamar Blue detection reagent provides a simple, rapid, reliable and safe method for cell proliferation and cytotoxicity detection, which is suitable for high-throughput detection experiments. The main component of the detection reagent is a redox indicator. In the oxidized stateProduct IntroductionAlamar Blue detection reagent provides a simple, rapid, reliable and safe method for cell proliferation and cytotoxicity detection, which is suitable for high-throughput detection experiments. The main component of the detection reagent is a redox indicator. In the oxidized state, it appears purple-blue and non-fluorescent, while in the reduced state, it turns into a reduction product with pink or red fluorescence, with an absorption peak of 530-560nm and an emission peak of 590nm.In the process of cell proliferation, the ratios of NADPH/NADP, FADH/FAD, FMNH/FMN and NADH/NAD in the cell increase and are in a reducing environment. The dye taken into the cell is reduced by these metabolic intermediates and cytochromes and then released outside the cell and dissolved in the culture medium, changing the culture medium from non-fluorescent indigo blue to fluorescent pink. Finally, use an ordinary spectrophotometer or fluorophotometer for detection, and the absorbance and fluorescence intensity are proportional to the number of active cells.Instructions1. Add 10µl of detection reagent to 100µl of cell suspension, and incubate in a cell incubator for 2-6 hours. The color of the medium changes from indigo blue to pink and you can proceed to the next step.2. It is recommended to use a fluorescence microplate reader for detection, the excitation light wavelength is between 530-560 nm, the emission light wavelength is 590 nm, and the relative fluorescence unit (RFU) is recorded.3. Draw a standard curve or cell growth curve: the ordinate (Y axis) is the relative fluorescence unit (RFU); the abscissa (X axis) is the cell number or time point or drug concentration.Precautions1. The appropriate density of cells can increase the detection sensitivity. For 96-well plates, we recommend seeding 100 microliters of cells per well. The cell concentration range is: 100-10,000/well for adherent cells, 2,000-50,000/well for suspension cells, and medium as a blank control. For 384-well plates, the cell concentration and seeding volume are both halved.2. The whole process should be aseptic operation, because microbial contaminants can also reduce the detection reagents and affect the experimental results.3. Pay attention to the concentration of inoculated cells and the incubation time after adding detection reagents. If the cell concentration is too high or the incubation time is too long, it will cause a secondary reduction reaction, resulting in colorlessness and disappearance of fluorescence.4. When incubating, avoid light.5. This product can use fluorescence or spectrophotometric detection, but the sensitivity of fluorescence is high, and the experimental error is small. Fluorescence detection is recommended... Read More | DescriptionIt contains a set of seven different homogeneous palladium catalysts, useful for rapid screening of catalysis conditions. It is in sampler format with individual components packaged for multiple experiments and mini scale-up. The cost of the kit is less than the total cost of individual DescriptionIt contains a set of seven different homogeneous palladium catalysts, useful for rapid screening of catalysis conditions. It is in sampler format with individual components packaged for multiple experiments and mini scale-up. The cost of the kit is less than the total cost of individual components.Catalysis Screening Kits... Read More | V669947 Component 50T Storage V669947A Buffer GL 15 mL RT V669947B Buffer GW1 (concentrate) 13 mL RT V669947C Buffer GW2 (concentrate) 15 mL RT V669947D Buffer RE 10 mL RT V669947E Proteinase K 12.5 mg RT V669947F Proteinase K Storage Buffer 1.25 mL RT V669947G Spin Columns RS with Collection Tubes V669947 Component 50T Storage V669947A Buffer GL 15 mL RT V669947B Buffer GW1 (concentrate) 13 mL RT V669947C Buffer GW2 (concentrate) 15 mL RT V669947D Buffer RE 10 mL RT V669947E Proteinase K 12.5 mg RT V669947F Proteinase K Storage Buffer 1.25 mL RT V669947G Spin Columns RS with Collection Tubes 50 RT V669947H RNase-Free Centrifuge Tubes (1.5 mL) 50 RTProductsThis kit is suitable for the extraction of viral RNA and DNA from fresh or frozen plasma, serum and cell-free body fluids. It is easy to operate as it does not require the use of organic solvents such as phenol and chloroform for extraction. The kit uses a unique buffer system to enable efficient and specific binding of viral nucleic acids in lysate to silica gel centrifugal adsorption columns. Inhibitors of PCR and enzyme reactions as well as residual impurities can be efficiently removed in a two-step effective rinsing step, and finally high purity viral nucleic acids can be obtained by using a low-salt buffer or water for elution. The purified viral nucleic acid is free of protein, nuclease and other impurities, and can be used directly in PCR, RT-PCR, Real-Time PCR, blotting experiments and so on.Self-contained reagent: anhydrous ethanol.Pre-experiment and Important Notes1. Add 1.25ml Proteinase K Storage Buffer to Proteinase K to dissolve it and store it at -20℃. Do not leave the prepared Proteinase K at room temperature for a long time, and avoid repeated freezing and thawing to avoid affecting its activity. Do not add Proteinase K directly into Buffer GL.2. Repeated freezing and thawing of the sample should be avoided, as this may result in smaller DNA fragments and a decrease in the amount of extracted DNA.3. Avoid repeated freezing and thawing of serum or plasma, which can lead to protein denaturation or precipitation, reducing the viral titer and thus affecting the yield of extracted viral nucleic acids.4. Anhydrous ethanol should be added to Buffer GW1 and Buffer GW2 according to the label instructions of the reagent bottle before first use.5. Check Buffer GL for crystallization or precipitation before use. If crystallization or precipitation occurs, redissolve Buffer GL in a water bath at 56℃.Procedure1. Take a 1.5 ml centrifuge tube (self-provided) and add 20 µl Proteinase K.2. Add 200 µl serum or plasma to the centrifuge tube. Add 200µl Buffer GL and vortex and shake for 15 seconds.Note: 1) Sample volume less than 200 µl can be made up by adding 0.9% NaCl (self-provided). 2) In order to ensure effective lysis of the sample, the sample needs to be mixed well with Buffer GL after adding Buffer GL.3. Incubate at 56°C for 15 minutes, centrifuge briefly, and collect the solution from the wall of the tube to the bottom of the tube.4. 250 µl of anhydrous ethanol was added, vortexed and shaken for 15 seconds, left at room temperature for 5 minutes, centrifuged briefly, and the solution on the wall of the tube was collected at the bottom of the tube.Note: If the ambient temperature exceeds 25°C, anhydrous ethanol should be used after pre-cooling on ice.5. Add the solution obtained in step 4 to the adsorbent column (RNase-Free Columns RS) that has been loaded into the collection tube, and if the solution cannot be added at one time, it can be transferred in several times. centrifuge the column at 12,000 rpm (~13,400 × g) for 1 min, pour off the waste liquid in the collection tube, and put the column back into the collection tube.6. Add 500 µl of Buffer GW1 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.7. Add 500 µl of Buffer GW2 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.Note: Step 7 can be repeated if further DNA purity is required.8. Add 500 µl of anhydrous ethanol to the adsorbent column and centrifuge at 12,000 rpm for 1 min. Pour off the waste liquid in the collection tube and put the adsorbent column back into the collection tube.9. Centrifuge at 12,000 rpm for 3 minutes and pour off the waste liquid in the collection tube. Leave the adsorption column at room temperature for several minutes to dry thoroughly.Note: The purpose of this step is the removal of residual ethanol from the adsorbent column; ethanol residue can interfere with subsequent enzymatic reactions (digestion, PCR, etc.).10. Place the adsorption column in a new collection tube (RNase-Free Centrifuge Tube), add 20-150 µl of Buffer RE or sterilized water overhanging the middle of the adsorption column membrane, leave it at room temperature for 2-5 minutes, and then centrifuge it at 12,000 rpm for 1 minute to collect the nucleic acid solution.Note: 1) If the downstream experiment is sensitive to pH or EDTA, you can use sterilized water for elution. The pH of the eluent has a great influence on the elution efficiency, if water is used as the eluent it should be ensured that its pH is 7.0-8.5 (the pH of water can be adjusted to this range with NaOH), and the elution efficiency is not high when the pH is lower than 7.0.(2) For long-term storage, please store the DNA solution at -20℃ and the RNA solution at -70℃.3) If the final concentration of DNA/RNA is to be increased, the DNA/RNA eluate obtained in step 10 can be re-spiked onto the adsorbent membrane and step 10 repeated... Read More |